EVALUATION OF IN VITRO ACARICIDAL ACTIVITY OF Cyclea peltata (PADAL)
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Date
2022-02-25
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COLLEGE OF VETERINARY AND ANIMAL SCIENCES MANNUTHY, THRISSUR, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY
Abstract
The present study was conducted to identify the acaricidal activity of potent fraction/ extracts of Cyclea peltata (Padal) against Rhipicephalus (Boophilus) annulatus. The whole plant was collected locally, dried, powdered and extracted. The aqueous extract of C. peltata was prepared by using hot extraction process and the methanolic extract was done in Soxhlet extraction apparatus and concentrated under rotary vacuum evaporator. The methanolic extract fractionated using solvents n-hexane, dichloromethane, n-butanol and water in increasing order of polarity to acquire different fractions. Qualitative Phytochemical screening of all the extracts as well as GCMS and FTIR analysis of the most potent fraction was performed. The acaricidal activity was determined by evaluating various entomological parameters such as mean egg mass, index of egg laying, per cent adult mortality and per cent inhibition of fecundity. The most potent fraction (n-hexane) was screened for its safety in HepG2, hepatocellular carcinoma cell lines by MTT assay and the IC50 was calculated. The cells were exposed to AO/EB and JC-1 staining to assess the mechanism of cytotoxicity of the extract. The phytochemical analysis unveiled the presence of steroids, alkaloids, glycosides, flavonoids, terpenoids, saponins in aqueous extract and alkaloids, phenols, tannins, flavonoids, terpenoids, saponins in methanolic extract. The fractions of C. peltata showed the presence alkaloids, phenols, tannins, flavonoids, terpenoids, saponins. There was no significant difference in the weight of ticks exposed to different extracts/fractions of C. peltata compared to control. The maximum reduction in egg mass was observed with n-hexane fraction with values of 0.002 ± 0.00 g, followed by methanolic and aqueous extracts whereas, there was minimum reduction in egg mass when ticks were treated with dichloromethane, n-butanol and water fractions of C. peltata compared to control. The index of egg laying was minimum when ticks were treated with n-hexane fraction with a value 0.044 ± 0.15 g whereas it was maximum when ticks were treated with n-butanol fraction of C. peltata (0.359±0.15 g). The aqueous, methanolic and n-hexane fractions of C. peltata produced 100 per cent adult mortality whereas the other fractions produced nearly 80 per adult mortality. The n-hexane fraction of C. peltata showed almost 90 per cent inhibition of fecundity while minimum inhibition was seen with n-butanol treatment.From the results, it was evident that out of all the extracts and fractions considered, n-hexane fraction exhibited lower mean egg mass and induced very early adult tick mortality with 91.38 per cent inhibition of fecundity. Hence, n-hexane fraction of C. peltata was found to be most potent fraction and was selected for in vitro evaluation of safety in HepG2 cell lines. GC-MS analysis of the n-hexane potent fraction divulged the presence of alkaloids, flavonoids, phenols, terpenoids and saponins. FTIR analysisrevealed the presence of Decanol, octyl-beta-d-glucopyranoside, 9-octadecenoic acid, cis androsterone, 1,7-heptanediol, ethyl undecanoate etc,. A dose dependent cytotoxicity was shown by n-hexane fraction in HepG2 cells with decrease in cell viability. Based on the per cent inhibition, the IC50 of n-hexane fraction was calculated as 8 µg/mL. Acridine
orange/Ethidium bromide staining (AO/EB) ascertained that most of the cells exposed to n-hexane fraction of C. peltata showed were in apoptotic stages and JC-1 staining elucidated that, the mitochondrial membrane potential declined in these cells.
From the present study, it was evident that n-hexane fraction of C. peltataexhibited potent acaricidal activity against R. (B.) annulatus than other extracts/ fractions with effective cytotoxic property in HepG2 cells and can be used as a lead for the
development of ectoparasiticide.